Structural Basis for the Mechanism and Substrate Specificity of Glycocyamine Kinase, a Phosphagen Kinase Family Member
Structural Basis for the Mechanism and Substrate Specificity of Glycocyamine Kinase, a Phosphagen Kinase Family Member
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DOI：
10.1021/bi9020988
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发表时间：
2010-03-09
期刊：
影响因子：
2.9
通讯作者：
Herzberg, Osnat
中科院分区：
文献类型：
--
作者：
Lim, Kap;Pullalarevu, Sadhana;Herzberg, Osnat
糖胞胺激酶（Glycocyamine kinase， GK）是磷酸激酶家族的一员，它催化磷酸糖胞胺n -磷酸基向ADP的Mg2+依赖性可逆磷酸基转移，生成糖胞胺和ATP。在一些多中心生物中，这种反应有助于维持细胞的能量稳态，以应对高且可变的能量周转。GK前面的地幔蠕虫namalycasttis sp.是异二聚体，具有两个同源的多肽链，α和β，由一个共同的前mrna通过互斥的n端交替外显子衍生而来。GK β的n端外显子编码的肽序列不同，比GK α的n端外显子编码的肽长16个氨基酸。重组GK α β和GK β β的晶体结构分别在2.6和2.4埃分辨率下进行了测定。此外，在2.3埃分辨率下，用过渡态模拟物测定了GK β β的结构。与序列同源性一致，file GK亚基采用与其他已知结构的磷酸腺苷激酶（同二聚体肌酸激酶（CK）和单体精氨酸激酶（AK））相同的总体折叠。与CK一样，GK的n端介导二聚体界面。在异二聚体和同二聚体的GK形式中，两个n末端的构象都是不对称的，这种不对称与之前报道的来自几种生物的同二聚体的ck不同。GK α - β的整个多肽链在结构上是明确的，β亚基的较长n端锚定在二聚体界面上。在GK β β中，一个亚基的24个n端残基和第二个亚基的11个n端残基是无序的。这一观察结果与一项建议是一致的，即一旦异源二聚体作为酶的生理形式出现，参与界面形成的GK α - β氨基酸就会得到优化。结果，同型二聚体界面（单独的α链或单独的β链）被破坏。在未结合状态下，GK表现出与无配体CK或AK类似的开放构象。在结合过渡态类似物后，GK的两个亚基都经历相同的闭合运动，从而闭合过渡态类似物，而CK的过渡态类似物只占据一个亚基，并进行结构域闭合。GK、CK和AK在结合状态下的活性位点环境揭示了底物特异性的结构决定因素。尽管gk二聚体的两个活性位点具有相同的结合，但n端的构象不对称仍被保留。因此，先前提出的CK的结构不对称与负协同性之间的耦合在GK的情况下不被支持。
Glycocyamine kinase (GK), a member of the phosphagen kinase family, catalyzes the Mg2+-dependent reversible phosphoryl group transfer of the N-phosphoryl group of phosphoglycocyamine to ADP to yield glycocyamine and ATP. This reaction helps to maintain the energy homeostasis of the cell in some multicenter organisms that encounter high and variable energy turnover. GK front the mantle worm Namalycastis sp. Is heterodimeric, With two homologous polypeptide chains, alpha and beta, derived from a common pre-mRNA by mutually exclusive N-terminal alternative exons. The N-terminal exon of GK beta encodes a peptide that is different in sequence and is 16 amino acids longer than that encoded by the N-terminal exon of GK alpha. The crystal Structures of recombinant GK alpha beta and GK beta beta from Namalycastis sp. were determined at 2.6 and 2.4 angstrom resolution, respectively. In addition, the structure of the GK beta beta was determined at 2.3 angstrom resolution in complex with a transition state analogue. Mg2+-ADP-NO3--glycocyamine Consistent with the sequence homology, file GK subunits adopt the same overall fold its that of other phospha-en kinases of known structure (the homodimeric creatine kinase (CK) and the monomeric arginine kinase (AK)). As with CK, the GK N-termini mediate the dimer interface. In both heterodimeric and homodimeric GK forms, the conformations of the two N-termini are asymmetric, and the asymmetry is different than that reported previously for the homodimeric CKs from several organisms. The entire polypeptide chains of GK alpha beta are structurally defined, and the longer N-terminus of the beta subunit is anchored at the dimer Interface. In GK beta beta the 24 N-terminal residues of one subunit and 11 N-terminal residues of the second subunit are disordered. This observation is consistent with a proposal that the GK alpha beta amino acids involved ill the interface formation were optimized once a heterodimer emerged as the physiological form of the enzyme. As a consequence, the homodimer Interface (either solely alpha or solely beta chains) has been corrupted. Ill the unbound state, GK exhibits an open conformation analogous to that observed with ligand-free CK or AK. Upon binding the transition state analogue, both subunits of GK undergo the same Closure motion that, clasps the transition State analogue, In contrast to the transition state analogue complexes of CK, where the corresponding transition State analogue Occupies only one subunit, which undergoes domain closure. The active site environments of the GK, CK, and AK at the bound states reveal the structural determinants Of Substrate Specificity. Despite the equivalent binding ill both active sites of the G K dimer, the conformational asymmetry of the N-termini is retained. Thus, the coupling between the structural asymmetry and negative cooperativity previously proposed for CK is not Supported ill the case of GK.